Bovine Pancreatic Trypsin Inhibitor (BPTI) is one of the smallest and simplest globular proteins. BPTI's sole function is to bind to and inhibit proteolytic enzymes like trypsin. BPTI contains both helical and sheet regions, as well as three disulfide bonds, which help to stabilize the tertiary structure of the molecule.

With three S-S bridges in 58 residues, BPTI is one of the stablest proteins known. It is quite inert to denaturants like urea and exhibits thermal denaturation below 100 C only in very acidic solutions; the half-point for reversible denaturation is about 80 C at pH 2.1 (Figure 6.23). But if only one of the disulfide bonds (that between cysteine residues 14 and 38) has been reduced and carboxymethylated, the midpoint is decreased to 59 C.

When all the disulfide bonds in BPTI are reduced, the protein is unfolded at room temperature. Yet upon reoxidation (re-forming the S-S bonds), native protein with the three correct disulfide pairings is efficiently formed. This re-formation is not what would be expected by chance. Suppose a BPTI molecule has been reduced, yielding 6 cysteine residues, and we now randomly reoxidize the SH groups. The first SH group to pick a partner will have 5 choices, the second group 3, and the last only 1, so there are 5 x 3, or 15, equally probable combinations. Thus, we would expect only about 7% of reduced BPTI to refold successfully by chance. But many studies of this and other proteins containing disulfide bonds indicate that correct pairing is regained if sufficient time is allowed. This finding must mean that it is the preferred folding of the protein that places the SH groups in position for correct pairing. The corollary of this statement is that the S-S bridges are not themselves essential for correct refolding. They do, however, contribute to the stability of the structure once it is folded. A molecule containing S-S bridges has a smaller number of conformations available in the unfolded form than does a comparable protein without the bridges. Consequently it shows a smaller entropy gain on unfolding and is therefore stabilized.

See also: -Helix

-Sheet